title: The involvement of β-amyrin 28-oxidase (CYP716A52v2) in oleanane-type ginsenoside biosynthesis in Panax ginseng.
abstract: Panax species are the most popular medicinal herbs. The root of these plants contains pharmacologically active triterpene saponins, also known as ginsenosides, compounds that are divided into dammarane- and oleanane-type triterpenes. Two CYP716A subfamily genes (CYP716A47 and CYP716A53v2) were recently characterized, encoding an enzyme catalyzing the hydroxylation of dammarane-type triterpenes in Panax ginseng. Herein, we report that one CYP716A subfamily gene (CYP716A52v2) isolated from P. ginseng encodes a β-amyrin 28-oxidase, which is suggested to modify β-amyrin into oleanolic acid, a precursor of an oleanane-type saponin (mainly ginsenoside Ro) in P. ginseng. The ectopic expression of both PNY1 and CYP716A52v2 in recombinant yeast resulted in erythrodiol and oleanolic acid production, respectively. In vitro enzymatic activity assays biochemically confirmed that CYP716A52v2 catalyzed the oxidation of β-amyrin to produce oleanolic acid, and the chemical structure of the oleanolic acid product was confirmed using gas chromatography-mass spectrometry (GC/MS). Transgenic P. ginseng plants were generated via Agrobacterium tumefaciens-mediated transformation: the overexpression of CYP716A52v2 greatly increased the content of oleanane-type ginsenoside (ginsenoside Ro), whereas RNA interference against CYP716A52v2 markedly reduced it. Furthermore, the levels of other dammarene-type ginsenosides were not affected in these transgenic lines. These results indicate that CYP716A52v2 is a β-amyrin 28-oxidase that plays a key role in the biosynthesis of oleanane-type triterpenes in P. ginseng.
authors.affiliation: Department of Forest Resources, College of Forest and Environmental Sciences, Kangwon National University, Chunchon 200-701, Republic of Korea.
authors.forename: Jung-Yeon
authors.initials: JY
authors.lastname: Han
authors.forename: Min-Jun
authors.initials: MJ
authors.lastname: Kim
authors.forename: Yong-Wook
authors.initials: YW
authors.lastname: Ban
authors.forename: Hwan-Su
authors.initials: HS
authors.lastname: Hwang
authors.forename: Yong-Eui
authors.initials: YE
authors.lastname: Choi
mesh_terms: D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D036145:Ginsenosides / Q000096:biosynthesis*; D009828:Oleanolic Acid / Q000031:analogs & derivatives* / Q000096:biosynthesis; D005894:Panax / Q000201:enzymology* / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D036145:Ginsenosides; D010940:Plant Proteins; C413246:oleanane; C064378:ginsenoside Ro; D009828:Oleanolic Acid; D003577:Cytochrome P-450 Enzyme System
keywords: CYP716A52v2; Cyt P450; Ginsenoside; Ginsenoside Ro; Oleanolic acid